1. Field of the Invention
The present invention relates to a serum-free culture medium used for the culture of animal cells.
2. Description of the Prior Art
In producing materials from animal cells, the selection of the culture medium in which the cells are cultured is very important since the production cost depends upon the price of the culture medium as well as the purifying costs of the final products.
Previously, a culture medium has been used containing about 10% of animal sera added to the basal synthetic culture medium for culturing animal cells.
However, the addition of sera increased the cost of the culture medium. Not only that, but the purifying cost of the product was likewise raised by the enriched protein contained in the sera.
Thus, a serum-free culture medium has been developed containing no sera. The serum-free culture medium is in general use at present for producing various materials. It is also known that various cell growth supplements can be added to a serum free culture medium. Representative examples are insulin, ethanol amine, sodium selenite, transferrin, albumin, lipoprotein, cytokine, and the like. Those which may raise the cost of the culture medium and the purifying cost of products include protein growth supplements. It is, therefore, indispensable to develop a serum-free culture medium containing little or essentially no protein.
The one which is comparatively expensive is transferrin among the growth supplements which are added to the serum-free culture medium.
Transferrin is considered to function as a growth supplement which is efficient in supplying cells with iron ions. (Newest Medical Science, Vol. 40, Growth Promotion by Transferrin" by Kimura and Ozawa, pages 554-559, 1985, and "Cell Growth Factor", pages 152-155, compiled by Japan Association of Tissue Culture Science, 1980).
The following recycling model of transferrin is asserted by Kogo, et al.
Iron-saturated transferrin is taken into cells by receptor-mediated endocytosis. After iron ions are released within the cells, transferrin is again secreted outside of the cells. ("Anemia by Iron Membrane Permeability Troubles" by Niitsu and Kogo, Separate Volume 3, Course of Medical Science, Medical Topics, page 124, 1988).
The above suggests that transferrin continues to supply cells with iron ions repeatedly.
On the contrary, Mather and Sato have reported that, in the serum-free culture medium containing F12 as its basal synthetic culture medium, the growth promotion by transferrin was replaced by highly concentrated iron ions. (J. P. Mather and G. H. Sato, Experiment Cell Research, Vol. 120, pages 191-200, 1979).
Further, Kovar and Franek report that, in the serum-free culture medium for mouse hybridoma composed of RPMI 1640 as the basal synthetic culture medium, the addition of a high concentration (500 .mu.M) of ferric citrate makes it possible to culture the hybridoma in non-protein serum-free culture. (Biotechnology Letters, Vol 9, pages 259-264, 1987).
However, it is known that a high concentration of iron ions is not desirable for cells. A serum-free culture medium should be developed capable of efficiently supplying cells with iron at lower concentrations.